This invention relates to a pharmaceutical composition comprising as an active ingredient a 2-piperidone compound which has antitumor activity and is useful as a medicament such as an antitumor agent.
Although studies have been widely made on antitumor agents efficacious on solid tumors, there are only few antitumor agents having low toxicity. The present inventors investigated antitumor agents efficacious on solid tumors and, as a result, found that certain 2-piperidone compounds are efficacious on solid tumors while showing low toxicity, thus completing the present invention.
4,6-Diphenyl-5-nitro-2-piperidone has been known as an intermediate for the synthesis of 3-aminopiperidine derivatives (U.S. Pat. No. 5,232,929). However, no pharmacological activity of this compound has been known. Also, 5-nitro-6-phenyl-1-(2-phenylethyl)-2-piperidone has been known (Synthesis, 615-616 (1976)) but its pharmacological activity is also unknown.
An object of the present invention is to provide 2-piperidone compounds which have an activity of inhibiting the proliferation of solid tumor cells and are useful as excellent antitumor agents.
The present invention relates to a 2-piperidone compound represented by the following formula (I) or a pharmaceutically acceptable salt thereof: 
wherein
R1 represents xe2x80x94(CH2)nR1a {wherein n is an integer of from 0 to 5, and R1a represents amino, lower alkylamino, di(lower alkyl)amino, substituted or unsubstituted aryl, or a substituted or unsubstituted heterocyclic group}, and
R2 and R3 independently represent lower alkyl which may be substituted by lower alkoxycarboyl; lower alkenyl, aralkyl or lower alkynyl which may be substituted by substituted or unsubstituted aryl or a substituted or unsubstituted heterocyclic group; substituted or unsubstituted aryl; or a substituted or unsubstituted heterocyclic group. Among the compounds of the formula (I), the compounds wherein R1a represents a substituted or unsubstituted heterocyclic group and R2 and R3 independently represent substituted or unsubstituted aryl or pharmaceutically acceptable salts thereof are preferable, and the compounds wherein R1a represents a heterocyclic group, and R2 and R3 independently represent substituted aryl or pharmaceutically acceptable salts thereof are further preferable. The present invention also relates to a pharmaceutical composition which comprises as an active ingredient the aforementioned 2-piperidone compound or a pharmaceutically acceptable salt thereof. The present invention further relates to an antitumor agent which comprises as an active ingredient the aforementioned 2-piperidone compound or a pharmaceutically acceptable salt thereof.
The present invention further relates to a pharmaceutical composition which comprises the 2-piperidone compound of the formula (I) or a pharmaceutically acceptable salt thereof; an antitumor agent which comprises the 2-piperidone compound of the formula (I) or a pharmaceutically acceptable salt thereof; a method for preventing or treating a patient having tumor, which comprises administering to the patient an effective amount of any one of the 2-piperidone compounds of the formula (I) or a pharmaceutically acceptable salt thereof; and use of any one of the 2-piperidone compounds of the formula (I) or a pharmaceutically acceptable salt thereof for the production of a pharmaceutical composition which is effective in preventing or treating a patient having tumor; use of any one of the 2-piperidone compounds of the formula (I) or a pharmaceutically acceptable salt thereof for the prevention or treatment of a patient having tumor; and a pharmaceutical composition which comprises a pharmaceutically acceptable carrier and an effective amount of any one of the 2-piperidone compounds of the formula (I) or a pharmaceutically acceptable salt thereof in a pharmaceutically acceptable dosage form.
Hereinafter, the compounds represented by the above formula (I) will be referred to as Compounds (I), and the same will apply to compounds represented by other formula numbers.
In the definition of each group given in the formula (I), the term xe2x80x9carylxe2x80x9d stands for a mono- to tricycle of 3- to 7-membered carbon rings wherein at least one of the rings is an aromatic ring. Examples thereof include phenyl, naphthyl, anthracenyl, tetrahydronaphthyl, indanyl and phenanthrenyl.
Examples of the heterocyclic group include azepinyl, benzimidazolyl, benzofurazanyl, benzopyranyl, benzothiopyranyl, benzofuryl, benzothiazolyl, benzothiadiazolyl, benzothienyl, benzoxazolyl, 1,4-benzodioxanyl, chromanyl, cinnolinyl, dihydrobenzofuryl, dihydrobenzothienyl, dihydrobenzothiopyranyl, furyl, imidazolidinyl, imidazolyl, imidazothiazolyl, indolinyl, indolyl, isochromanyl, isoindolyl, 1,3-dioxolanyl, 1,3-dithiolanyl, isoxazolyl, isoquinolyl, isothiazolyl, isothiazolidinyl, morpholinyl, naphthyridinyl, oxadiazolyl, oxazolyl, 2-oxoazepinyl, 2-oxopiperazinyl, 2-oxopyrrolidinyl, piperidyl, piperazinyl, pyridyl, pyridyl N-oxide, pyrazinyl, pyrazolinyl, pyrazolyl, pyrimidinyl, pyrrolidinyl, pyrrolyl, quinazolinyl, quinolinyl, quinoxalinyl, tetrahydrofuryl, tetrahydroisoquinolyl, tetrahydroquinolyl, tetrahydropyranyl, tetrazolyl, thiadiazolyl, thiazolyl, thiazolinyl, thienofuryl, thienothienyl, thienyl, triazolyl and pyridonyl.
Examples of the aralkyl include C7-20 aralkyl such as benzyl, phenethyl, benzhydryl, naphthylmethyl and trityl.
The lower alkyl and the lower alkyl moieties in the lower alkylamino, di(lower alkyl)amino and lower alkoxycarbonyl include linear, branched or cyclic C1-10 alkyl such as methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, pentyl, isopentyl, hexyl, heptyl, octyl, nonyl, decyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl and adamantyl.
The lower alkenyl includes linear, branched or cyclic C2-10 alkenyl such as vinyl, allyl, crotyl, 1-propenyl, prenyl, isopropenyl, 2-methyl-2-butenyl, pentenyl, hexenyl, heptenyl, 2,6-dimethyl-5-heptenyl, cyclobutenyl, cyclopentenyl, and cyclohexenyl.
The lower alkynyl includes linear or branched C2-10 alkynyl such as ethynyl, propynyl, butynyl, pentynyl, hexynyl, heptynyl, and octynyl.
Substituents on the aryl or heterocyclic group may be the same or different 1 to 3 substituents, and examples thereof include hydroxy; halogen; nitro; amino; carboxy; lower alkyl optionally substituted by 1 to 3 halogen, lower alkoxycarbonyl or hydroxy; lower alkoxy optionally substituted by 1 to 3 halogen or lower alkoxy; lower alkoxycarbonyl; lower alkylthio; lower alkenyl optionally substituted by 1 to 3 lower alkoxy, lower alkoxycarbonyl or a heterocyclic group; lower alkynyl; aryl optionally substituted by 1 to 3 halogen atoms; aryloxy; aryloxy(lower alkyl); aroyloxy; lower alkylamino optionally substituted by a heterocyclic group; hydroxyamino; formyl; lower alkanoyl; lower alkanoyloxy; amino substituted by lower alkanoyloxy or lower alkanoyl; di(lower alkyl)amino; di(lower alkyl)aminocarbonyloxy; lower alkanoylamino; lower alkylsulfonylamino; lower alkoxycarbonylamino; aralkylamino; sulfamylamino (aminosulfonylamino); ureido (carbamoylamino); cyano; aralkyl; aralkyloxy; arylsulfonyl; a heterocyclic group; a heterocyclic group-carbonyloxy; camphanoyloxy; methylenedioxy; ethylenedioxy; B(OH)2 and SO3H. The term xe2x80x9chalogenxe2x80x9d stands for fluorine, chlorine, bromine and iodine atoms. The aralkyl and the aralkyl moieties in the aralkylamino and the aralkyloxy are each as defined above. The lower alkyl and the lower alkyl moieties in the lower alkoxy, lower alkoxycarbonyl, aryloxy(lower alkyl), lower alkanoyl, lower alkylthio, lower alkylamino, lower alkanoyloxy, di(lower alkyl)amino, di(lower alkyl)aminocarbonyloxy, lower alkanoylamino, lower alkylsulfonylamino and lower alkoxycarbonylamino are each as defined above. The aryl and the aryl moieties in the aryloxy, aryloxy(lower alkyl), arylsulfonyl and aroyloxy are each as defined above. The heterocyclic group and the heterocyclic group moiety in the heterocyclic group-carbonyloxy are each as defined above. The lower alkenyl and the lower alkynyl are each as defined above.
Examples of the pharmaceutically acceptable salt of Compounds (I) include pharmaceutically acceptable acid addition salts, metal salts, ammonium salts, organic amine addition salts and amino acid addition salts thereof. Examples of the acid addition salt include inorganic acid addition salts such as hydrochlorides, hydrobromides, sulfates, phosphates and nitrates, and organic acid addition salts such as formates, acetates, propionates, benzoates, maleates, fumarates, succinates, tartrates, citrates, oxalates, methanesulfonates, p-toluenesulfonates, aspartates and glutamates. Examples of the metal salt include alkali metal salts such as lithium salts, sodium salts and potassium salts, alkaline earth metal salts such as magnesium salts and calcium salts, aluminum salts, and zinc salts. Examples of the ammonium salt include ammonium salts and tetramethylammonium salts. Examples of the organic amine addition salt include addition salts with morpholine and piperidine. Examples of the amino acid addition salt include addition salts with glycine, phenylalanine, glutamic acid and lysine.
Next, a process for producing Compounds (I) will be explained.
In the following process, when a group defined therein changes under the employed conditions or the group is inappropriate for carrying out the process, the object compound can be obtained by using the method of introducing and eliminating protective group(s) commonly used in the field of synthetic organic chemistry [see, for example, xe2x80x9cProtective Groups in Organic Synthesisxe2x80x9d, T. W. Greene, John Wiley and Sons, Inc. (1981)]. If necessary, the order of the reaction steps of, for example, introducing substituents may be appropriately changed.
Production Process
Compound (I) can be synthesized starting with Compound (II), which are known compounds or which can easily produce by a known process, via Compound (III) by, for example, the following steps. 
In the formula, R1, R2 and R3 are each as defined above.
(Step 1)
Compound (III) can be synthesized by reacting Compound (II) with 1 to 100 equivalents of nitromethane in an inert solvent (e.g., acetonitrile, dimethylformamide) in the presence of 0.01 to 10 equivalents of a base [e.g., 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU)] in accordance with a method reported in Synthesis, 226 (1984). The reaction is usually carried out at xe2x88x9230 to 100xc2x0 C. and completed within 1 to 72 hours. In this process, nitromethane can be employed also as the solvent.
(Step 2)
Compound (I) can be synthesized by reacting Compound (III) in a solvent (e.g., ethanol, methanol) with Compound (IV) represented by the following formula:
R3xe2x80x94CHOxe2x80x83xe2x80x83(IV)
wherein R3 is as defined above;
and Compound (V) represented by the following formula:
R1xe2x80x94NH2xe2x80x83xe2x80x83(V)
wherein R1 is as defined above. Usually, Compound (IV) and Compound (V) are used each in an amount of 1 to 5 equivalents based upon that of Compound (III). Also, an acid addition salt (e.g., acetate) of Compound (V) can be used instead of Compound (V). The reaction is usually carried out at 0 to 100xc2x0 C. and completed within 1 to 72 hours.
In the production of Compound (I), the conversion of the functional groups R1, R2 and R3 may be carried out by using a method for converting functional groups commonly employed in synthetic organic chemistry [see, for example, xe2x80x9cComprehensive Organic Transformationsxe2x80x9d, R. C. Larock (1989)].
In the above production process, the product can be isolated and purified by using an appropriately combination of techniques commonly employed in organic synthesis (e.g., filtration, extraction, washing, drying, concentration, crystallization, various types of chromatography).
Some of Compounds (I) can exist in the form of various stereoisomers such as enantiomers or diastereomers. All of the possible isomers and mixtures thereof including the above-mentioned ones fall within the scope of the present invention.
Compounds (I) and their pharmaceutically acceptable salts may exist in the form of adducts with water or various solvents. The above adducts can also be used as the treating agents according to the present invention.
Tables 1 to 6 show the structures and physical data of typical examples of Compound (I).
(wherein, R2a, R2b and R2c, and R3a, R3b and R3c respectively mean the substituents of the substituted phenyl groups as R2 and R3)
Next, the pharmacological activities of Compound (I) will be illustrated by reference to Test Examples.
Proliferation Inhibition Test on Human Colonic Cancer DLD-1 Cell
On a 96-well microtiter plate (Nunc #167008), 1,000/well of human colonic cancer DLD-1 cells were supplied, which were then pre-incubated in RPMI1640 medium containing 5% or 10% fetal calf serum (FCS) in a 5% carbon dioxide gas incubator at 37xc2x0 C. for 24 hours. Subsequently, a 10 mmol/L solution of each test compound in dimethylsulfoxide (DMSO) was diluted with the incubation medium and then further diluted 3-fold stepwise. The incubation was performed for additional 72 hours. After the completion of the incubation, the medium was discarded followed by the addition of 50 xcexcl/well of a solution of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (Sigma, hereinafter referred to simply as MTT) in the incubation medium (final concentration: 1 mg/ml). After maintaining the plate at 37xc2x0 C. in a 5% carbon dioxide gas incubator for 4 hours, the MTT solution was discarded and 150 xcexcl/well of DMSO was added. After completely dissolving the MTT-formazane crystals by vigorously stirring with the use of a plate mixer, the difference in the absorbances at 550 nM and 630 nM was measured by using a microplate reader SPECTRAmax 250 (Wako Pure Chemical Industries, Ltd.) The 50% inhibitory concentration (IC50) showing the proliferation inhibitory activity was calculated by using 4-parameter logistic curves of the included software SOFTmaxPRO.
Anti-tumor Activity Test on Human Colonic Cancer DLD-1 Solid Tumor Transplanted in Nude Mouse
Fragments (2 mmxc3x972 mm) were excised from well-proliferating parts of human colonic cancer cell DLD-1 tumor masses which had been subcultured in male nude mice (BALB/c-nu/nu mice, Clea Japan). Then, these fragments were subcutaneously transplanted into the abdominal region of 7-week-old male nude mice by using trocars. When the tumor volume (see, Formula-1) attained 50 to 70 mm3, the mice were divided into groups each having 5 animals. Next, each test compound was dissolved in physiological saline containing polyoxyethyelne sorbitan monooleate and the resulting solution was intraperitoneally administered to the mice twice a day continuously for 10 days. The value T/C (%) of each test compound was determined by measuring the tumor volume before the administration (V0) and that after the administration (V) and calculating the ratio (V/V0), as Formula-2 shows. The results are given in Table 8.
Tumor volume (mm3)={major diameter (mm)xc3x97(minor diameter (mm))2}xc3x971/2xe2x80x83xe2x80x83Formula-1
T/C (%)=((V/V0 of test group)/(V/V0 of control group))xc3x97100.xe2x80x83xe2x80x83Formula-2 
Proliferation Inhibition Test on Human Pancreatic Cancer MIA-PaCa2 Cell
On a 96-well microtiter plate (Nunc #167008), 2,000/well of human pancreatic cancer MIA-PaCa2 cells were supplied, which were then preincubated in RPMI1640 medium containing 10% fetal calf serum (FCS) in a 5% carbon dioxide gas incubator at 37xc2x0 C. for 24 hours. Subsequently, a 10 mmol/L solution of each test compound in dimethyl sulfoxide (DMSO) was diluted with the incubation medium and then further diluted 3-fold stepwise. The incubation was performed for additional 72 hours. After the completion of the incubation, the medium was discarded, followed by the addition of 50 xcexcl/well of a solution of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (Sigma, hereinafter referred to simply as MTT) in the incubation medium (final concentration: 1 mg/ml). After maintaining the plate at 37xc2x0 C. in a 5% carbon dioxide gas incubator for 4 hours, the MTT solution was discarded and 150 xcexcl/well of DMSO was added. After completely dissolving the MTT-formazane crystals by vigorously stirring with the use of a plate mixer, the difference in the absorbances at 550 nM and 630 nM was measured by using a microplate reader SPECTRAmax 250 (Wako Pure Chemical Industries, Ltd.). The 50% inhibitory concentration (IC50) showing the proliferation inhibitory activity was calculated by using 4-parameter logistic curves of the included software SOFTmaxPRO.
The above Compound (I) have an effect of inhibiting the proliferation of human colonic cancer DLD-1 cells and human pancreatic cancer MIA-PaCa2 cells and, therefore, are useful as the active ingredient of medicaments, preferably antitumor agents. The first embodiment of the pharmaceutical composition of the present invention is characterized by containing as the active ingredient substances selected from the group consisting of Compound (I), pharmaceutically acceptable salts thereof, and hydrates and solvates of the same. The pharmaceutical compositions of the present invention are useful as antitumor agents in treating non-solid cancers such as leukemia, malignant lymphoma and myeloma and solid cancers such as gastric cancer, esophageal cancer, intestinal cancer, rectum cancer, pancreatic cancer, hepatic cancer, renal cancer, bladder cancer, pulmonary cancer, uterus cancer, ovarian cancer, mammary cancer, prostatic cancer, skin cancer and brain tumor.
For the medicaments of the present invention, the above-mentioned substances as such may be administered as the active ingredient. However, it is generally preferable to administer them in the form of pharmaceutical compositions comprising the above substances together with one or more pharmaceutical additives. These pharmaceutical compositions can be produced by using methods which are well known or have been conventionally employed in the field of pharmaceutics. The medicaments of the present invention in the form of such pharmaceutical compositions may contain one or more other pharmaceutically active ingredients. The medicaments of the present invention are applicable to mammals including humans.
The medicaments of the present invention may be administered via an arbitrary route without restriction, and the most suitable administration route may be selected from oral and parenteral routes depending on the purpose of treatment and/or prevention. Examples of the parenteral administration route include tracheobronchial, rectal, subcutaneous, intramuscular and intravenous routes. Examples of the preparation appropriate for oral administration include tablets, granules, fine granules, powders, syrups, solutions, capsules and suspensions, while examples of the preparation appropriate for parenteral administration include injections, drips, inhalants, sprays, suppositories, transdermal absorption preparations and mucosal absorption preparations.
To prepare liquid preparations appropriate for oral administration, pharmaceutical additives such as water; saccharides (e.g., sucrose, sorbitol, fructose); glycols (e.g., polyethylene glycol, propylene glycol); oils (e.g., sesame oil, olive oil, soybean oil); preservatives (e.g., p-hydroxybenzoates); and the like can be used. To prepare solid preparations such as capsules, tablets or granules, excipients (e.g., lactose, glucose, sucrose, mannitol); disintegrating agents (e.g., starch, sodium alginate); lubricating agents (e.g., magnesium stearate, talc); binders (e.g., polyvinyl alcohol, hydroxypropylcellulose, gelatin); surfactants (e.g., fatty acid esters); and plasticizers (e.g., glycerol); and the like may be used.
Among the preparations appropriate for parenteral administration, those for intravascular administration (e.g., injections, drips) may be prepared preferably with the use of aqueous media isotonic to human blood. For example, injections may be prepared with the use of aqueous media selected from salt solutions, glucose solutions and saline/glucose solution mixtures in accordance with a conventional manner by using appropriate adjuvants to give solutions, suspensions or dispersions. Suppositories for rectal administration may be prepared by using carriers such as cacao fat, hydrogenated fats or hydrogenated carboxylic acids. Sprays may be prepared by using carriers which can promote the dispersion of the above-mentioned substances serving as the active ingredient in the form of fine particles and the absorption thereof without irritating human oral cavity or respiratory mucosa. As such a carrier, lactose, glycerol, or the like may be used. These preparations may be in the form of aerosols, dry powders, etc. depending on the properties of the above-mentioned substances serving as the active ingredient as well as the properties of the carriers selected. To produce preparations for parenteral administration, one or more pharmaceutical additives selected from diluents, flavors, preservatives, excipients, disintegrating agents, lubricating agents, binders, surfactants, plasticizers, and the like may be used. The dosage forms and production processes of the medicaments of the present invention are not restricted to those described above in detail.
The dose and administration frequency of the medicaments of the present invention are not particularly restricted but can be appropriately determined depending on various factors, for example, the type of the above-described substance as the active ingredient, the type of the cancer to be treated, the administration route, the age and body weight of the patient, and the conditions and severity of the disease. For example, about 0.01 to 500 mg/kg/day may be administered to an adult at a frequency of once to 5 times in a day or once in several days to several weeks, though neither the dose nor the administration frequency is restricted thereto. The medicament of the present invention can be used together with other antitumor agents. It is generally favorable to combine the medicament of the present invention with several antitumor agents differing in the functional mechanism.
Hereinafter, Examples and Production Examples of the present invention will be given.
Tablet
A tablet of the following composition is prepared in a conventional manner:
Capsule
A capsule of the following composition is prepared in a conventional manner:
The above components are mixed and packed into a gelatin capsule.
Injection
An injection of the following composition is prepared in a conventional manner: